Rheumatoid arthritis (RA) is a chronic inflammatory disease affecting upwards of 1% of the general population. The KRN murine arthritis model shares many disease characteristics with RA, including joint inflammation and eventual destruction of the synovial joints. This disease affects primarily distal joints, leaving other joints unaffected. In this model, mice develop arthritis by about one month of age. The autoreactive T and B cells recognize the autoantigen, glucose 6-phosphate isomerase (GPI). Anti-GPI antibodies alone transfer arthritis to native recipient mice. Using this model, our laboratory has made several novel findings, including, establishing a critical role for neutrophils, showing the rapid localization (6 min) of anti-GPI to the affected joints using micro positron emission tomography (mPET), and revealing that the anti-GPI B cell response initiates in the joint draining lymph nodes. We have also recently observed that mice lacking, FcR, TNF, mast cells, and neutrophils are resistant to arthritis, and the anti-GPI antibodies do not localize to their distal joints. In this application, we propose to study three different aspects of the initiation of arthritis. In Aim I, we propose to identify the pathophysiological mechanisms involved in the initial cascade of events in the joints that result in arthritis, with a focus on the role of the neutrophil. These studies will involve various knockout and reconstituted mice and the localization of the antibodies will be followed by mPET. In Aim II, we will study the immune physiology of the joint. These studies are based on our new observation that the local lymph nodes draining the limbs is where the anti-GPI B cells are first stimulated. We will test the affect of increased GPI levels and examine joint and lymph node APCs for their ability to stimulate anti-GPI T cells. We also propose to develop a new model in which we knock-in HEL, (soluble or cytoplasmic) into the collagen II locus. These mice will express HEL only in the joints, and will permit us in a definitive manner to ascertain when, how, and where is a joint specific antigen accessible to the immune system. In Aim III, we propose to study B and T cell tolerance to GPI using the KRN TCRtg, an anti-GPI immunoglobulin tg mouse, and an antigen-free mouse. Overall, from these proposed studies, we will obtain new insights into the critical role of the neutrophil in the initiation of the arthritis, how an arthritogenic autoimmune response initiates in the local lymph nodes, and how does self-tolerance fail.